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EVALUATION OF THE EFFECT OF PROTEIN OPRF OF PSEUDOMONAS AERUGINOSA AND ATOX FOR THE MATURATION OF DENDRITIC CELLS IN MICE IN VITRO


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Abstract

There are multiple experimental and clinical evidences (data) of the successful using of the cellular vaccines based on the DC in immunotherapy of the oncological and infectious diseases. Objective: To study the morphological features and immunophenotype of DC, generated from BALB/c mice bone marrow progenitor cells, under the influence of recombinant OprF и aTox Pseudomonas aeruginosa antigens. Materials and methods. Preparation: 25 μg of OprF, 50 μg of aTox sorbed by 75 μg of aluminum hydroxide. The recombinant protein preparation was mixed in equal weight fractions with an aluminum hydroxide gel, diluted in phosphate buffered saline (PBS), and sorbed for 12 hours at 4 °C. Dendritic cells (DC) were obtained from bone marrow cells of BALB/c mice when cultured with recombinant GM-CSF and IL-4 (Biosource, USA) for 6 days. OprF and aTox of P. aeruginosa (15 μl/ml) and commercial TNF-α (20 ng/ml, Biosource, USA) were used as the inducer of maturation of DC. Immunophenotype of the DC was estimated by using of the flow cytometry method. Results. We reviled that morphological features of the DC when used OprF/aTox antigens and their combination with aluminium hydroxid as inductor of DC maturation, have no sufficiently differences. Test preparations caused a decrease in the number of immature cells (CD34), an increase in the cell population with adhesion markers (CD38), antigen presentation of MHCII, costimulatory CD80 / CD86 molecules and a terminal differentiation molecule (CD83). Conclusions. The candidate Pseudomonas aeruginosa vaccine based on its recombinant OprF and aTox proteins OprF и aTox induces the maturation of the DC generated from mice bone marrow cells. The obtained data expand the possibilities for developing new sources of the inducers of DC maturation, and can also serve as a criterion for evaluating the effectiveness of the developed vaccine against Pseudomonas aeruginosa.


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